QTL for spot blotch resistance in bread wheat line Saar co-locate to the biotrophic disease resistance loci Lr34 and Lr46

Spot blotch caused by Bipolaris sorokiniana is a major disease of wheat in warm and humid wheat growing regions of the world including south Asian countries such as India, Nepal and Bangladesh. The CIMMYT bread wheat line Saar which carries the leaf tip necrosis (LTN)-associated rust resistance genes Lr34 and Lr46 has exhibited a low level of spot blotch disease in field trials conducted in Asia and South America. One hundred and fourteen recombinant inbred lines (RILs) of Avocet (Susceptible) × Saar, were evaluated along with parents in two dates of sowing in India for 3 years (2007–2008 to 2009–2010) to identify quantitative trait loci (QTL) associated with spot blotch resistance, and to determine the potential association of Lr34 and Lr46 with resistance to this disease. Lr34 was found to constitute the main locus for spot blotch resistance, and explained as much as 55 % of the phenotypic variation in the mean disease data across the six environments. Based on the large effect, the spot blotch resistance at this locus has been given the gene designation Sb1. Two further, minor QTL were detected in the sub-population of RILs not containing Lr34. The first of these was located about 40 cM distal to Lr34 on 7DS, and the other corresponded to Lr46 on 1BL. A major implication for wheat breeding is that Lr34 and Lr46, which are widely used in wheat breeding to improve resistance to rust diseases and powdery mildew, also have a beneficial effect on spot blotch.     

Enrichment of adeno-associated virus serotype 5 full capsids by anion exchange chromatography with dual salt elution gradients

Adeno-associated virus-based gene therapies have demonstrated substantial therapeutic benefit for the treatment of genetic disorders. In manufacturing processes, viral capsids are produced with and without the encapsidated gene of interest. Capsids devoid of the gene of interest, or “empty” capsids, represent a product-related impurity. As a result, a robust and scalable method to enrich full capsids is crucial to provide patients with as much potentially active product as possible. Anion exchange chromatography has emerged as a highly utilized method for full capsid enrichment across many serotypes due to its ease of use, robustness, and scalability. However, achieving sufficient resolution between the full and empty capsids is not trivial. In this work, anion exchange chromatography was used to achieve empty and full capsid resolution for adeno-associated virus serotype 5. A salt gradient screen of multiple salts with varied valency and Hofmeister series properties was performed to determine optimal peak resolution and aggregate reduction. Dual salt effects were evaluated on the same product and process attributes to identify any synergies with the use of mixed ion gradients. The modified process provided as high as ≥75% AAV5 full capsids (≥3-fold enrichment based on the percent full in the feed stream) with near baseline separation of empty capsids and achieved an overall vector genome step yield of >65%.     

Knockout of 5‐lipoxygenase prevents dexamethasone‐induced tau pathology in 3xTg mice

Emerging evidence suggests that dysregulation stress hormones, such as glucocorticoids, in aged persons put them at a higher risk to develop Alzheimer's disease (AD). However, the mechanisms underlying such vulnerability remain to be unraveled. Pharmacologic inhibition of 5‐lipoxygenase (5LO), an active player in AD pathogenesis whose protein level increases with aging in the human, has been shown to blunt glucocorticoid‐mediated amyloid β (Ab) formation in vitro. In this article, we investigated the role of this pathway in modulating the development of the corticosteroid‐dependent AD‐like phenotype in the triple transgenic mice (3xTg). Dexamethasone was administered for 1 week to 3xTg or 3xTg genetically deficient for 5LO (3xTg/5LO^?/?) mice, and its effect on memory, amyloid‐β and tau levels, and metabolism assessed. At the end of the treatment, we observed that dexamethasone did not induce changes in behavior. Compared with controls, treated mice did not show significant alterations in brain soluble Aβ levels. While total tau protein levels were unmodified in all groups, we found that dexamethasone significantly increased tau phosphorylation at S396, as recognized by the antibody PHF‐13, which was specifically associated with an increase in the GSK3β activity. Additionally, dexamethasone‐treated mice had a significant increase in the tau insoluble fraction and reduction in the postsynaptic protein PDS‐95. By contrast, these modifications were blunted in the 3xTg/5LO^?/? mice. Our findings highlight the functional role that 5LO plays in stress‐induced AD tau pathology and support the hypothesis that pharmacologic inhibition of this enzyme could be a useful tool for individuals with this risk factor.     

High copper levels in the medium improves shoot bud differentiation and elongation from the cultured cotyledons of <Emphasis Type="Italic">Capsicum annuum</Emphasis> L.

The effect of copper sulphate on differentiation and elongation of shoot buds from cotyledonary explants of Capsicum annuum L. cv X-235 was investigated. Shoot buds were induced on medium supplemented with 22.2 μM BAP and 14.7 μM PAA. Elongation of shoot buds was obtained on MS medium containing 13.3 μM BAP + 0.58 μM GA₃. Both shoot induction and elongation media were supplemented with different levels of CuSO₄ (0–5 μM). The levels of CuSO₄ in the induction as well as elongation medium highly influenced the shoot bud formation and their subsequent elongation. Highest number of shoot buds per explant was obtained when the concentration of CuSO₄ was increased 30 times to the normal MS level. Shoot buds formation frequency i.e., the number of shoots formed per explant was increased two fold as compared to those formed on control. Elongation both in terms of percentage and length of shoots was better than that on control. Healthy elongated shoots were rooted on MS medium supplemented with 5.7 μM IAA. Rooted plantlets were transferred to field conditions.     

Regulation of cardiac sarcoplasmic reticulum (Ca2+ + Mg2+)-ATPase

Summary The two high affinity calcium binding sites of the cardiac (Ca²⁺ + Mg²⁺)-ATPase have been identified with the use of Eu³⁺. Eu³⁺ competes for the two high affinity calcium sites on the enzyme. With the use of laser-pulsed fluorescent spectroscopy, the environment of the two sites appear to be heterogeneous and contain different numbers of H₂O molecules coordinated to the ion. The ion appears to be occluded even further in the presence of ATP. Using non-radiative energy transfer studies, we were able to estimate the distance between the two Ca²⁺ sites to be between 9.4 to 10.2 A in the presence of ATP. Finally, from the assumption that the calcium site must contain four carboxylic side chains to provide the 6–8 ligands needed to coordinate calcium, and based on our recently published data, we predict the peptidic backbone of the two sites.     

Cloning, expression, and characterization of the human mitochondrial beta-ketoacyl synthase. Complementation of the yeast CEM1 knock-out strain

A human beta-ketoacyl synthase implicated in a mitochondrial pathway for fatty acid synthesis has been identified, cloned, expressed, and characterized. Sequence analysis indicates that the protein is more closely related to freestanding counterparts found in prokaryotes and chloroplasts than it is to the beta-ketoacyl synthase domain of the human cytosolic fatty acid synthase. The full-length nuclear-encoded 459-residue protein includes an N-terminal sequence element of approximately 38 residues that functions as a mitochondrial targeting sequence. The enzyme can elongate acyl-chains containing 2-14 carbon atoms with malonyl moieties attached in thioester linkage to the human mitochondrial acyl carrier protein and is able to restore growth to the respiratory-deficient yeast mutant cem1 that lacks the endogenous mitochondrial beta-ketoacyl synthase and exhibits lowered lipoic acid levels. To date, four components of a putative type II mitochondrial fatty acid synthase pathway have been identified in humans: acyl carrier protein, malonyl transferase, beta-ketoacyl synthase, and enoyl reductase. The substrate specificity and complementation data for the beta-ketoacyl synthase suggest that, as in plants and fungi, in humans this pathway may play an important role in the generation of octanoyl-acyl carrier protein, the lipoic acid precursor, as well as longer chain fatty acids that are required for optimal mitochondrial function.     

Gender comparisons between unilateral and bilateral landings

The increased number of women participating in sports has led to a higher knee injury rate in women compared with men. Among these injuries, those occurring to the ACL are commonly observed during landing maneuvers. The purpose of this study was to determine gender differences in landing strategies during unilateral and bilateral landings. Sixteen male and 17 female recreational athletes were recruited to perform unilateral and bilateral landings from a raised platform, scaled to match their individual jumping abilities. Three-dimensional kinematics and kinetics of the dominant leg were calculated during the landing phase and reported as initial ground contact angle, ranges of motion (ROM) and peak moments. Lower extremity energy absorption was also calculated for the duration of the landing phase. Results showed that gender differences were only observed in sagittal plane hip and knee ROM, potentially due to the use of a relative drop height versus the commonly used absolute drop height. Unilateral landings were characterized by significant differences in hip and knee kinematics that have been linked to increased injury risk and would best be classified as "stiff" landings. The ankle musculature was used more for impact absorption during unilateral landing, which required increased joint extension at touchdown and may increase injury risk during an unbalanced landing. In addition, there was only an 11% increase in total energy absorption during unilateral landings, suggesting that there was a substantial amount of passive energy transfer during unilateral landings.     

The formation of germtubes by candida albicans,when grown with Staphylococcus pyogene,Escherichia coli,Klebsiella pneumoniae,Lactobacilius acidophilus and Proteus vulgaris

The formation of germtubes by twelve clinical isolates of C. albicans was studied in human serum containing per millilitre 10³ to 10⁹ organisms as: Staphylococcus pyegene, Escherichia coli, Klebsiella pneumoniae, Lactobacillus acidophilus and Proteus vulgaris. All the five bacteria inhibited formation of germtubes by C. albicans at all concentrations and the percent germtube formed diminished with increasing concentration of the bacteria. Lactobacillus acidophilus inhibited the formation of germtubes maximally followed by Staphylococcus pyogene, Escherichia coli and Klebsiella pneumoniae. Proteus vulgaris in the concentrations of 10³ to 10⁷ bacteria per millilitre produced only insignificant inhibition of formation of germtubes by C. albicans. Since germtubes of C. albicans are invasive, it is suggested that inhibition of blastospo-regermtube transformation may be significantly responsible for prevention of infection by C. albicans by coexisting bacterial flora.     

Nitric oxide-mediated modulation of synaptic activity by astrocytic P2Y receptors

We investigated the mechanism of synaptic suppression by P2Y receptors in mixed hippocampal cultures wherein networked neurons exhibit synchronized Ca²⁺ oscillations (SCO) due to spontaneous glutamatergic synaptic transmission. Pharmacological studies suggested that SCO suppression was mediated by P2Y2/P2Y4 receptors. Immunostaining studies and characterization of ATP/UTP-stimulated Ca²⁺ responses in solitary neurons and astrocytes revealed that the SCO attenuation was effectuated by astrocytes. We demonstrate that nitric oxide released from activated astrocytes causes synaptic suppression by inhibiting neurotransmitter release. Physiological concentrations of ATP and UTP evoked NO production in astrocytes. SCO suppression was considerably diminished by removal of extracellular NO by membrane-impermeable scavenger c-PTIO or by pretreatment of cells with nitric oxide synthase inhibitor L-NAME. The nitric oxide donor DETA/NO effectively suppressed the SCO. ATP/UTP inhibited KCl-induced exocytosis at presynaptic terminals in an NO-dependent manner. In the absence of exogenously added ATP/UTP, both the NO scavenger and NOS inhibitor enhanced the frequency of SCO, implying that astrocytes release NO during spontaneous synaptic activity and exert a suppressive effect. We report for the first time that under physiological conditions astrocytes use NO as a messenger molecule to modulate the synaptic strength in the networked neurons.